The single layer of endothelial cells that line the vascular lumen are nonthrombogenic and are unique in expressing VIII related von Willebrand factor. These cells are also known to express several other factors important to hemostasis/thrombosis including factor V, antithrombin III, plasminogen activator, and thrombospondin. Endothelial cells will replicate to a limited extent in culture and continue to express these differentiated functions. Intracellular factor VIII related antigen (VIIIR:Ag) is organized into distinct foci, and in plasma its associated platelet aggregating activity is directly related to its multimeric size. The importance of the multimeric assembly of VIIIR:Ag to its function is indicated by human genetic variants in which long bleeding times are associated with smaller VIIIR:Ag multimers. Other genetic variants which affect this factor differently suggest that more than one genetic determinant may be involved. The main objective of this project is to construct somatic cell hybrids of endothelial cells and certain heterologous cells which are perpetual in culture and may be permissive for the expression of differentiated functions related to hemostasis. These hybrids would be used to determine which chromosomes carry the genes for factor VIII related antigen and other endothelial cell specific functions by correlating their expression with the presence of biochemical chromosome markers. Cultured endothelial cells will also be used to investigate how the large biologically active multimers of von Willebrand factor are assembled and glycosylated. Correlation of the expression of factor VIII related antigen with other differentiated functions related to hemostasis/thrombosis will be investigated with regard to intracellular localization of antigens, to rate of expression as a function of time in culture, to sensitivity to glycosylation blocking agents, and to suppression by hybridization with nonpermissive heterologous cells.